Martin Wiedmann, Dr. med. vet., PhD
Listeria monocytogenes causes both animal and human infections. While human listeriosis is a relatively rare human foodborne disease, L. monocytogenes is a considerable concern for the food industry and for public health due to the severity of the disease, which includes 20-30% case mortality rates and hospitalization rates for afflicted humans of >90%. This presentation and proceedings article will highlight key aspects of the ecology and transmission of L. monocytogenes, with a focus on linkages between animal infections and human listeriosis.
Listeria monocytogenes and Listeriosis
Listeria monocytogenes is a facultative intracellular gram-positive bacterium associated with potentially serious invasive diseases in humans and in more than 40 animal species.1 Ruminants (cattle, sheep, and goats) seem to be more commonly diagnosed with listeriosis than other animal species. Infection in these hosts can result in abortion, septicemia, meningitis and encephalitis as well as other less severe manifestations, including diarrhea and skin infections. Among humans, immunocompromised persons, pregnant women, the elderly and neonates are particularly at risk of listerial infections. While the majority of human clinical infections occur as sporadic cases, human listeriosis can also occur in large epidemics. The vast majority (99%) of human Listeria infections is thought to be foodborne.2 The frequency of clinical human listeriosis in most developed countries is estimated to range between 2 to 15 cases/million population with case mortality rates between 13 and 34%.3 A study published in 1999 estimated that a total of 2,500 human listeriosis infections, including 500 deaths, occurred in the US annually.2 Thus, although foodborne listeriosis cases are less common than many other foodborne diseases (e.g., those caused by E. coli O157:H7, Campylobacter jejuni, or Salmonella), they represent the second most common known cause of fatal human foodborne infections in the US, next only to Salmonella infections.
Since L. monocytogenes represents a foodborne pathogen, a considerable numbers of studies and surveys on the presence of this organism in many raw and ready-to-eat foods (RTE) have been published. While the prevalence of L. monocytogenes varies tremendously by study, type of food surveyed, and other factors, this organism has been isolated from many different foods, occasionally at fairly high prevalences (>5-10%).3,4 Most recent surveys of RTE foods in developed countries show L. monocytogenes prevalences considerably below 5% however.5,6 The ability of L. monocytogenes to survive and proliferate well in foods stored at refrigeration temperatures makes this organism a particular concern for the RTE food industry, since low initial contamination levels (possibly even at <1 cfu/25g) may increase to numbers that could present a human health hazard if products are subject to extended refrigerated storage. Consequently, humans appear to be commonly exposed to L. monocytogenes by food ingestion. Based on the 2001 FDA/USDA Draft Listeria monocytogenes risk assessment,7 the average US consumer is likely to occasionally (possibly once a year) consume as many as 106 to 109 cfu of L. monocytogenes in a single serving. In spite of this apparent occasional high exposure, only 2,500 human listeriosis cases occur annually in the US. These data indicate that even exposure to high levels of this organism may not result in human disease, even when one considers that the at-risk population for human listeriosis (immunocompromised people, pregnant women etc.) represents only a fraction of the total US population.
Transmission and Ecology of Listeria monocytogenes
The pathogenesis of foodborne infectious diseases can involve complex interactions between a bacterial pathogen, a variety of environments and one or multiple host species. The ability of bacterial cells to survive and compete in a variety of environments plays a key role in the pathogenesis and transmission of many foodborne diseases. In a simplified model, transmission patterns of L. monocytogenes as well as of other foodborne pathogens may encompass all or some of the following steps and environments: 1) bacterial survival in the environment and in animal feeds (e.g., L. monocytogenes and pathogenic E. coli are common environmental contaminants); 2) bacterial survival inside non-vertebrate hosts (e.g., protozoans); 3) establishment of clinical or subclinical infections or carrier states in food animals; 4) shedding of the organism into animal products used for human consumption or secondary contamination of animal products; 5) bacterial survival and/or multiplication in non-host environments under food processing and distribution conditions; and 6) infection of human hosts, including survival of gastric passage and establishment of enteric or systemic infections.
Human L. monocytogenes infections are most commonly traced back to contaminated RTE foods that allow the growth of L. monocytogenes and that are stored for extended times under refrigeration temperature, thus allowing growth of this pathogen. L. monocytogenes is effectively inactivated by the heat treatments generally used for production of RTE foods. While it thus appears that most L. monocytogenes contamination of RTE foods occurs as post-processing contamination, either in the processing plant, at retail, or the home, the primary origin (and reservoir) of the L. monocytogenes found in these environments is usually not apparent.
Subtype Analysis of L. monocytogenes
As L. monocytogenes is commonly found in many different environments, understanding transmission of this foodborne pathogen requires the ability to differentiate subtypes and strains of this pathogen. While serotyping has been traditionally used for subtype characterization of bacterial pathogens, including many foodborne pathogens, DNA-based subtyping methods generally provide for better discriminations. Common molecular subtyping methods used for L. monocytogenes include ribotyping as well as Pulsed Field Gel Electrophoresis (PFGE). PFGE is the most common subtyping method used for foodborne pathogens and its use as part of national and international foodborne disease surveillance systems (i.e., PulseNet) has considerably extended our ability to detect foodborne disease outbreaks and their sources.
Transmission of Listeria monocytogenes
Survival of L. monocytogenes outside human and animal hosts plays an important role in the transmission of this organism in the food system. While it is conceivable that L. monocytogenes can enter the food chain at many and possibly even at nearly any point, agricultural environments and food processing plant environments may be of particular importance as sources for the introduction of L. monocytogenes into the food system. The specific studies conducted by my research groups and described below indicate that 1) L. monocytogenes, including those types responsible for human disease, is more common in bovine farms than any other environments and appears to efficiently multiply in bovine farms, indicating that the bovine farm environment may represent a reservoir for L. monocytogenes; 2) L. monocytogenes, including the types responsible for human disease, can persist in processing plants and retail establishments for months to years and these persistent types appear to be a main source of finished product contamination. In addition, data by our group have also shown that more human listeriosis cases than previously assumed may represent small outbreaks. While it is clear that direct transmission from farm animals to humans is rare (unlike for pathogens such as Salmonella and E. coli); farms and farm environments may still play an important role as primary (even though not necessarily as direct) sources of human pathogenic L. monocytogenes.
L. monocytogenes Transmission and Ecology in Animal-based Agricultural Systems
Listeriosis in farm animals is often linked to consumption of contaminated silage. In addition, animal listeriosis cases sometimes occur in animals that are not fed silage and environmental sources have been speculated to be responsible for at least some of these cases. The agricultural environment thus may serve not only as an important source for contamination of silage, but may also be a direct source of animal infection in some cases. The role of contaminated raw animal-based agricultural products (e.g., milk, meat) as a direct source of L. monocytogenes contamination of RTE food products is likely to be minimal since commercially applied heat treatments generally kill L. monocytogenes effectively enough to provide an appropriate margin of safety. While infected animal and contaminated agricultural environments appear to rarely be a direct cause of human infections, there have been cases where a direct link has been observed. Specifically, animal sources can play an important role in animal-derived food products that are not processed before consumption (e.g., raw milk). In addition, manure from infected or shedding animals may represent a source of food contamination. For example, an outbreak involving 42 human cases in Nova Scotia in 1981 was linked to the consumption of coleslaw. This coleslaw was produced from cabbage harvested from fields fertilized with untreated sheep manure that had been obtained from a farm with a history of ovine listeriosis.8
To better understand the transmission of L. monocytogenes in farm environments, we conducted a case-control study involving 24 case farms with at least one recent case of listeriosis and 28 matched control farms with no listeriosis cases.9 A total of 528 fecal, 516 feed and 1012 environmental soil and water samples were cultured for L. monocytogenes. While the overall L. monocytogenes prevalence was similar in cattle case (24.4%) and control (20.1%) farms, small ruminant (goat and sheep) farms showed a significantly higher prevalence (p<0.0001) in case (32.9%) than control farms (5.9%). Interestingly, while 20.1% of the 2056 farm samples collected were positive for L. monocytogenes, the L. monocytogenes prevalence among 1805 soil, water, and other environmental samples from urban and pristine environments (i.e., locations with little direct animal and human contact, such as state parks, national forests, etc.) also collected by our group during the same time period and in the same region (New York state) was considerably lower (1.3% and 7.3% for pristine and urban environments, respectively).10 EcoRI ribotyping of clinical (n=17) and farm isolates (n=413) differentiated 51 ribotypes. L. monocytogenes ribotypes isolated from clinical cases and fecal samples were more frequent in environmental than in feed samples, indicating that infected animals may contribute to L. monocytogenes dispersal into the farm environment. Our data indicate that 1) the epidemiology and transmission of L. monocytogenes differs between small ruminant and cattle farms; 2) cattle contribute to amplification and dispersal of L. monocytogenes into the farm environment, 3) the bovine farm ecosystem maintains a high prevalence of L. monocytogenes, including subtypes linked to human listeriosis cases and outbreaks, and 4) L. monocytogenes subtypes may differ in their abilities to infect animals and to survive in farm environments.
L. monocytogenes Transmission in Food Processing Environments
Increasing evidence indicates that the processing plant environment may be one of the most important sources of food product contamination with L. monocytogenes. We thus have focused considerable research efforts on developing a better understanding of the ecology of L. monocytogenes in food processing plants. In a pilot study using three smoked fish processing plants, we showed that specific L. monocytogenes subtypes persisted in the environment of a given processing plant.11 In addition to these persistent, plant-specific, subtypes, we also observed a considerable diversity of transient L. monocytogenes subtypes. Interestingly, the persistent L. monocytogenes subtypes were the major cause of finished product contamination. Through a collaborative research project on the ecology of L. monocytogenes in Mexican-style cheese processing plants, we also showed a similar pattern of strain distribution in dairy processing plants, including persistence of plant-specific subtypes over time. In one of the three plants studied, the persistent L. monocytogenes subtype was also responsible for finished product contamination.12 These findings are consistent with other reports that used bacterial subtyping methods to show the persistence of specific L. monocytogenes subtypes in a variety of food processing environments including those for smoked fish, poultry, meat and dairy foods.13-16 Our results indicate that control and elimination of persistent L. monocytogenes subtypes in processing plants may allow for considerable reduction of finished product contamination. Molecular subtyping provides an important tool to track in-plant sources and spread of bacterial contaminants in order to better control finished product contamination by foodborne pathogens.
L. monocytogenes Transmission and Ecology in Retail Environments
While considerable information on L. monocytogenes contamination patterns in food processing plants is available, our understanding of L. monocytogenes contamination and transmission in retail operations is limited. We characterized 125 food, 40 environmental and 342 human clinical L. monocytogenes isolates collected in New York State from 1997 to 2002 using automated ribotyping.17 All environmental isolates were obtained from retail establishments and the majority of food isolates (105 isolates) were obtained from foods that were prepared or handled at retail. Overall, food and environmental isolates from 53 different retail establishments were characterized. The 125 food and 40 environmental isolates were differentiated into 29 and 10 ribotypes, respectively. For 16 retail establishments, we found evidence for persistence of one or more specific L. monocytogenes as indicated by isolation of the same EcoRI ribotype from food and/or environmental samples collected in a given establishment on different days. The human isolates were differentiated into 48 ribotypes. A total of 17 ribotypes found among the human clinical isolates were also found among the food and environmental isolates though. We thus conclude that L. monocytogenes, including subtypes that have been linked to human disease, can persist in retail environments. Implementation of Listeria control procedures in retail operations that process and handle products that permit the growth of L. monocytogenes are thus a critical component of a farm-to-table L. monocytogenes control program.
Epidemiology of Human Listeriosis
While large foodborne listeriosis outbreaks have been reported, the majority of reported human cases are generally classified as sporadic cases for which no source was identified. To gain a better understanding of the epidemiology of human listeriosis, we have conducted a survey of human L. monocytogenes isolates in New York state using automated EcoRI ribotyping and PFGE as subtyping techniques.18 A total of 34 ribotypes and 74 PFGE types were found among 131 human isolates. Using the scan statistic (P < 0.05), 9 clusters (31% cases) were identified by either ribotype or PFGE typing with 4 clusters (16% cases) identified using both subtyping methods. Two of the 10 clusters identified (13% cases) corresponded to epidemiologically supported multi-state listeriosis outbreaks. Based on these data, we concluded that while most human listeriosis cases have been considered sporadic, highly discriminatory molecular subtyping approaches indicate that a substantial number of case clusters occur. At least 13% and as many as 31% of the human listeriosis cases reported in New York State may represent single-source, multi-case clusters. Listeriosis reduction and control efforts thus need to include broad-based subtyping of human isolates and need to consider that a significant number of cases may represent outbreaks.
While direct transmission of L. monocytogenes from animals to humans is likely to occur only rarely (unlike for other foodborne pathogens such as Salmonella and E. coli O157:H7), due to its common high prevalence in farms and farm animals, particularly dairy cows, attention must be paid to minimize transmission of L. monocytogenes from animals to humans, particularly through raw milk and dairy products made from raw milk as well as possibly through fruits and vegetables exposed to raw manure.
1. Seeliger HPR. In: Listeriosis, 1961, p. 60. Hafner Publishing Co., Inc., New York;
2. Mead P, et al. Emerg Infect Dis 1999; 5:607;
3. Farber JM, Peterkin PI. Microbiol Rev 1991; 55:476;
4. Farber JM, Peterkin PI. In: Listeria, listeriosis, and food safety, 1999, p. 505 (Ryser ET. and Marth EH, Eds.) Marcel Dekker, Inc., New York;
5. Ryser ET. In: Listeria, listeriosis, and food safety. 1999, p. 411(Ryser ET. and Marth EH, Eds.) Marcel Dekker, Inc., New York;
6. Ryser ET. In: Listeria, listeriosis, and food safety, 1999. p 359 (Ryser ET. and Marth EH, Eds.) Marcel Dekker, Inc., New York;
7. Food and Drug Administration and US Department of Agriculture. 2001. Draft assessment of the relative risk to public health from foodborne Listeria monocytogenes among selected categories of Ready-to-Eat foods. USDA, FDA, Washington DC (http://www.foodsafety.gov/~dms/lmrisk.html);
8. Schlech WFI, et al. N Engl J Med 1983; 308:203;
9. Nightingale KK, et al. Appl. Environ. Microbiol 2004; 70 4458;
10. Sauders BD, et al. J Food Prot 2006; 69:93;
11. Norton D, et al. Appl Environ Microbiol 2001; 67:198;
12. Kabuki DY, et al. J Dairy Sci 2004; 87:2803;
13. Autio T, et al. Appl Environ Microbiol 1999; 65:150;
14. Lawrence LM, Gilmour A. Appl Environ Microbiol 1995;61:2139;
15. Nesbakken T, et al. Int J Food Microbiol 1996; 31:161;
16. Rorvik L, et al. Int J Food Microbiol 1995; 25:19;
17. Sauders BD, et al. J. Food. Prot 2004; 67:1417.
18. Sauders BD, et al. Emerg Infect Dis 2003; 9:672